The role of miR-146a/IRAK1/JNK1 pathway in mediating the effects of Yiqi Congming decoction on dry eye: A mechanistic study in rat models

ETHNOPHARMACOLOGICAL RELEVANCE: Yiqi Congming Decoction (YQCM) is a traditional Chinese medicine formula widely used as a complementary and alternative therapy for dry eye and other ophthalmic disorders. OBJECTIVE: This study aims to investigate the potential effects of YQCM on dry eye and to identify the active components responsible for its therapeutic efficacy using a rat model. MATERIALS AND METHODS: Using Ultra-High Performance Liquid Chromatography/Quadrupole Time-of-Flight Mass Spectrometry (HPLC-QTOF/MS), the chemical constituents of YQCM were identified. In vivo experiments demonstrated the protective effects of YQCM on the corneal barrier in a rat model of dry eye and determined the optimal therapeutic dose. YQCM and agomiR-146a were administered either individually or in combination to rat over 14 days, followed by evaluations of Schirmer I test (SⅠT) results, tear film breakup time (BUT), fluorescein staining (FL) levels, corneal epithelial cell inflammation, and apoptosis. Furthermore, the expression of proteins in the miR-146a/IRAK1/JNK1 pathway, as well as inflammation and apoptosis-related proteins, was examined to explore the mechanisms through which YQCM modulates inflammation and improves tear film stability. In vitro experiments employed serum containing YQCM, agomiR-146a, and the IRAK1 inhibitor (AZ1495) to further investigate the regulatory effects of YQCM on the miR-146a/IRAK1/JNK1 pathway and its impact on inflammation and apoptosis in human corneal epithelial cells (HCECs) under hypertonic conditions. RESULTS: In vivo experimental results demonstrated that YQCM significantly restored tear film stability. Treatment with varying doses of YQCM improved corneal epithelial damage in rats, with the medium dose exhibiting the most pronounced effect. YQCM increased the SⅠT and BUT levels, effectively reduced FL levels, and inhibited apoptosis and inflammatory damage in corneal epithelial cells. Additionally, scanning electron microscopy, hematoxylin-eosin (HE) staining, TUNEL assays, and Western blot (WB) and qPCR analyses revealed that YQCM significantly ameliorated corneal damage in dry eye rats and reduced the expression levels of MMP-9, TNF-α, IL-1β, Caspase-3, IL-6, and IFN-γ. Moreover, YQCM modulated the expression of proteins in the miR-146a/IRAK1/JNK1 pathway. In vitro experiments demonstrated that YQCM regulated the levels of miR-146a/IRAK1/JNK1 in HCECs under hypertonic conditions and enhanced cell viability by reducing the expression of MMP-9, TNF-α, IL-1β, Caspase-3, IL-6, and IFN-γ. Using Annexin V-FITC/PI double staining and other techniques, it was further confirmed that YQCM alleviated apoptosis in HCECs under hypertonic conditions. CONCLUSION: YQCM protects tear film stability in a rat model of dry eye by suppressing corneal epithelial inflammatory responses and reducing apoptosis through modulation of the miR-146a/IRAK1/JNK1 pathway.